During liver regeneration quiescent differentiated hepatocytes replicate to restore the hepatic mass. Hepatocyte replication in the regenerating liver has a high degree of synchrony and involves the transcription and posttranscriptional activation of transcriptional factors, protooncogenes and growth factors among many other genes. In work supported by this grant (years 25-29) we demonstrated that cytokines are involved in the initiation of liver regeneration and that signaling through Tumor Necrosis Receptor type 1 (TNFR-1) required for liver regeneration by activating a pathway that includes NKkappaB, IL-6 and STAT3. Work in liver cell cultures developed in this laboratory as well as with RelA/TNFR-1 double knockout mice demonstrated that signaling through TNFR-1 can be proliferative, apoptotic or cause the transcription of acute phase response genes. This work led to the conclusion that hepatocytes need to be primed by cytokines to fully respond to the growth factors HGF and TFGalpha. However, major questions remain regarding the cytokine-dependent and independent mechanisms that initiate regeneration as well as the nature of signaling interactions between hepatocytes and Kupffer cells which may be required for hepatocyte replication. Furthermore, little information is available about mechanisms that may limit cytokine expression as well as DNA replication in the regenerating liver. We have designed experiments to address 4 specific questions using carefully selected mouse models and hepatocyte cell cultures: a) whether blockage of NFkappaB activation in hepatocytes but not in Kupffer cells will interfere with liver regeneration; b) whether the blockage of the generation of Reactive Oxygen Species (ROS) inhibits hepatocyte DNA replication; c) whether LPS is responsible for TNF induction at the start of liver regeneration; d) whether 2 distinct negative feedback mechanisms acting on cytokine signaling and MAP kinase activity may respectively regulate the initiation and cell cycle progression phases of liver regeneration.